Clostridia are gram-positive, anaerobic, sporeforming bacteria with a fermentative metabolism. Clostridia are very effective producers of gas (H.sub.2 and CO.sub.2); their other fermentation products include butyric acid, acetic acid, butanol, ethanol, isopropanol, acetone and other organic acids and alcohols. Clostridia are also the principal agents of the anaerobic decomposition of proteins. The soil and water are the primary environment of clostridia, although they also may be present in poor-quality silage, feeds, fertilizers and contaminated foods, such as vegetables, fish, and crustaceans, meat, milk, and sweets.
Certain clostridia have been found to be pathogenic. Pathogenic clostridia are also normally found in the soil. They have little invasive power, but instead they produce a variety of highly toxic proteins known as exotoxins, and clostridial diseases are a result of such exotoxins. Examples of diseases caused by clostridia include botulism and other less hazardous food poisonings, which are specifically a result of the exotoxins formed in the food. Other severe clostridial diseases include tetanus and gas gangrene, resulting from wound infections. Some clostridial toxins, such as those responsible for botulism and tetanus, are potent inhibitors for nerve function. Others again, such as those causing gas gangrene, are enzymes. Such enzymes include lecithinase, hemolysin and a variety of different proteases.
In addition to being pathogenic, clostridia are also otherwise harmful. The shelf-life and utility of foods and their raw materials is often impaired by clostridial spoilage. Clostridia spoil not only food consumed fresh or employed as raw material in the food industry but also products of the food industry, such as products of the canning industry, processed meat, milk products, etc. The effects of clostridia on food have been described in W. C. Frazier, Food Microbiology, 2nd edition, 1967, McGraw-Hill Book Company, New York.
Clostridia may also present problems in food production. Particularly in the dairy industry, the adverse effects of food-borne clostridia are great. Hence one of the greatest problems attending cheese production is false fermentation caused by clostridia during the curing of cheese. Clostridia use for example available carbohydrates and lactic acid from normal lactic acid fermentation, producing therefrom butyric acid and gases, particularly H.sub.2 and CO.sub.2. Such butyric acid fermentation will completely spoil the cheese. On account of excessive gas formation, the texture of the cheese changes, the cheese swells too much and may even explode. The butyric acid formed in cheese has a very strong and unpleasant taste, and thus the cheese is not accepted for vending on account of the foul off-taste.
Various means are employed in an attempt to inhibit the growth of clostridia. Nitrate and nitrite in the form of various alkali salts are used to inhibit clostridial growth for example in meat and meat products and in certain cheese varieties. However, nitrates and nitrites are toxic at high concentrations, and their use as additives is not desired. In view of the consumer attitudes that are increasingly against the use of additives and the different food legislations in different countries, industry in general currently strives to find natural biopreservatives to replace chemical additives.
Heat treatment, such as sterilization of foods or their raw materials, has also been employed in an attempt to inhibit clostridial growth. Spores formed by clostridia are nevertheless thermostable, and in the worst case heat treatment can even have the opposite effect. In the dairy industry, problems are also presented by the fact that pasteurization of milk kills other acid-forming bacteria, and thus clostridia which do not normally grow very well at low temperatures and on the other hand do not compete well with other acidforming bacteria at higher temperatures have no competing organisms after the spores have germinated.
To avoid problems caused by clostridia, it is attempted to keep the clostridial spore content in milk as low as possible by completely prohibiting the use of silage or by using silage of very high quality only. Clostridial spores can also be bactofugated from milk prior to the use of the milk as such or as a raw material in the dairy industry. Bactofugation, i.e. removal of bacteria and/or spores by centrifugation, nevertheless has various effects on the quality of the final product and is also uneconomic.
To inhibit the growth of clostridia, the enzyme lysozyme has also been used in several branches of the food industry. The activity of lysozyme is not confined to harmful microbes only, but lysozyme also inhibits the activity and growth of any desired bacteria. Thus such a treatment also affects the quality of the final products and is not recommended for use.
Lactic acid bacteria are known to produce various antimicrobial compounds, such as organic acids, hydrogen peroxide, diacetyl and bacteriocins, and it has been attempted to use these as additives for instance to improve the shelf-life of food. Commercially available is the product Nisaplin (Aplin & Barrett), incorporating purified nisin produced by the bacterium Lactococcus lactis and exerting an effect against gram-positive bacteria. The use of nisin to control the growth of Clostridium botulinum spores in high moisture content cheese spreads is described in U.S. Pat. No. 4 584 199. U.S. Pat. No. 4 790 994 discloses the use of Pediococcus pentosaceus to inhibit the growth of psychrotrophic bacteria in milk products of e.g. the cottage cheese type. There is no reference to clostridia in this patent.
WO Patent publication 93/09676 relates to a method for preserving a food product by treating the product with an euhygienic bacterial strain competitively inhibiting the growth of pathogenic and spoilage organisms prior to packaging of the product. Euhygienic bacteria are defined as non-pathogenic and/or non-spoilage bacteria. Lactobacillus delbruckii and Hafnia alvei are stated as preferred. However, the effect of these species has not been shown in the publication.
European published application 344 786 (A2) discloses a bacterial preparation incorporating a Lactobacillus gasseri, Lactobacillus delbruckii and/or Bifidobacterium longum culture in a suitable medium, such as soy milk, which according to the publication can be used to inhibit clostridial growth. The publication indicates that the preparation can be used both therapeutically, to inhibit intestinal clostridial growth, and as a food preservative, in which case the preparation is used to increase the acidity of the product. However, the anticlostridial effect of said bacteria has not been demonstrated.
DE Offenlegungsschrift 31 25 797 (A1) discloses a dietetic preparation comprising Lactobacillus casei in a suitable culture medium, such as soy milk, which in accordance with the publication can be used to inhibit intestinal clostridial growth. The publication indicates that the preparation can, like other similar lactic acid bacteria preparations, also be used for prophylaxis for example in undergoing antibiotic therapy. However, the publication provides no test results of the anticlostridial effects of the dietetic preparation or the Lactobacillus casei culture incorporated therein, and there is no proof of the effectiveness of the invention. Neither is any other utility for Lactobacillus casei described or suggested in the publication.